Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.11.11 (AMPK)
 12,425 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The cAMP response element-binding protein (CREB) mediates transcriptional activation of genes in response to the cAMP signal transduction pathway. There are two different isoforms of CREB, which are generated by alternative RNA splicing. There is evidence that the two isoforms may have different biological activities. As the longer isoform (CREB341) contains a potential phosphorylation site that is not present in the shorter isoform (CREB327), we examined the possible differential phosphorylation of the two CREB isoforms. Recombinant CREB was prepared and used as substrate for phosphorylation by the cAMP-dependent protein kinase in vitro. Phosphopeptide mapping and mutagenesis studies demonstrated that CREB341 contains two sites, serine 133 and serine 98, that can be phosphorylated in vitro by the catalytic subunit of the cAMP-dependent protein kinase. In contrast, CREB327 contains only a single phosphorylation site at serine 119 (equivalent position to serine 133 in CREB341). A kinase titration experiment demonstrated that serine 98 of CREB341 was phosphorylated only at relatively high concentrations of the cAMP-dependent protein kinase. Transient transfection studies were used to test for any possible function of the phosphorylation of serine 98 of CREB341. These studies used GAL4-CREB fusion proteins. We found that mutation of serine 98 to alanine (which would block phosphorylation) has little or no effect on the ability of the CREB fusion protein to activate transcription. These findings suggest that differences in the biological activity of the two CREB isoforms are probably not mediated by differential phosphorylation by the cAMP-dependent protein kinase.
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PMID:Phosphorylation of cyclic adenosine 3',5'-monophosphate (cAMP) response element-binding protein isoforms by the cAMP-dependent protein kinase. 148 Jan 75

A mechanism by which voltage-sensitive Ca2+ channel (VSCC) activation triggers c-fos transcription has been characterized. Ca2+ influx through VSCCs stimulates phosphorylation of the transcription factor cAMP response element-binding protein (CREB) on serine 133 leading to an increase in the formation of transcription complexes that can elongate through a transcription pause site within the c-fos gene. Ca(2+)-stimulated CREB serine 133 phosphorylation is mediated by a Ca(2+)-activated kinase and is not dependent on the cAMP-dependent protein kinase (PKA). While necessary for c-fos transcriptional induction following VSCC opening, CREB serine 133 phosphorylation is not sufficient for transcriptional activation. A second, PKA-dependent event is required. Following induction, c-fos transcription is rapidly down-regulated. Dephosphorylation of CREB serine 133 parallels and likely mediates the transcriptional shut-off event. These results suggest that the phosphorylation and dephosphorylation of CREB controls its ability to regulate transcription in membrane-depolarized cells and that multiple pathways contribute to Ca(2+)-activated gene expression.
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PMID:L-type voltage-sensitive Ca2+ channel activation regulates c-fos transcription at multiple levels. 787 82

8-Chloro-cyclic AMP (8-Cl-cAMP) produces growth-inhibitory and differentiating activity in the promyelocytic leukemia cell line HL-60. Adriamycin (ADR)-resistant HL-60 (HL-60/AR) cells exhibit the multidrug-resistant phenotype but do not express the mdr1 gene product P-glycoprotein. To explore potential signaling processes that may be involved in this atypical form of drug resistance, 8-Cl-cAMP was used as a modulator of the cAMP second messenger signal transduction pathway. Treatment for 48 hr with a 10% inhibitory concentration of 8-Cl-cAMP potentiated ADR cytotoxicity 14-fold in HL-60/AR cells but not in the parental cell line. 8-Cl-cAMP was stable to hydrolysis in the medium after 48 hr and was present intracellularly predominantly as phosphorylated metabolites (70%) and the parent compound (30%). No difference occurred in ADR accumulation in HL-60/AR cells after treatment with 8-Cl-cAMP. Accompanying the 8-Cl-cAMP-mediated increase in ADR cytotoxicity in HL-60/AR cells was a reduction in the cytosolic type I cAMP-dependent protein kinase (PKA) and disappearance of the nuclear PKA holoenzyme. Coincident with these changes in drug-resistant cells was a marked reduction in the DNA-binding activity of the cAMP response element-binding protein to levels equivalent to those in sensitive cells. This effect appears to result from reduced phosphorylation of the cAMP response element-binding protein. These results suggest that the potentiation by 8-Cl-cAMP of ADR cytotoxicity in HL-60/AR cells occurs through down-regulation of nuclear type I PKA and cAMP response element-binding factors whose activities are regulated by PKA.
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PMID:Reversal of resistance to adriamycin by 8-chloro-cyclic AMP in adriamycin-resistant HL-60 leukemia cells is associated with reduction of type I cyclic AMP-dependent protein kinase and cyclic AMP response element-binding protein DNA-binding activities. 838 2

Activity-mediated gene expression is thought to play an important role in many forms of neuronal plasticities. We have used pentylenetetrazol-induced seizure that produces synchronous and sustained neuronal activity as a model to examine the mechanism(s) of gene activation. The transcription factor CREB (Ca2+/cAMP response element-binding protein) is thought to be necessary for long-term memory formation both in invertebrates and vertebrates. When phosphorylated on Ser133 either by cAMP-dependent protein kinase and/or Ca2+/calmodulin-dependent protein kinases, CREB increases transcription of genes containing the CRE (cAMP response element) sequence. Using an antibody that detects Ser133-phosphorylated CREB protein, we show that CREB phosphorylation is maximal between 3 and 8 min after the onset of seizure activity and declines slowly both in the hippocampus and the cortex. The total amount of CREB protein did not change at the time points examined. The increased phosphorylation of CREB protein is preceded by an increase in the amount of cAMP, suggestive of cAMP-dependent protein kinase activation, in the hippocampus and activation of Ca2+/calmodulin-dependent protein kinases in the cortex. Subsequent to CREB phosphorylation, the expression of the CRE-containing gene, c-fos, and the AP-1 complexes (heterodimers of Fos and Jun family members) is increased. These findings support the role of CREB-mediated gene expression in activity-dependent neuronal plasticities.
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PMID:Neuronal activity increases the phosphorylation of the transcription factor cAMP response element-binding protein (CREB) in rat hippocampus and cortex. 866 77

Transcription factor, cAMP response element-binding protein (CREB), which is phosphorylated by cAMP-dependent kinase via an increase in cAMP, and regulates gene transcription by binding to the cAMP response element (CRE) on target genes. We examined age-dependent alterations in the DNA-binding activity of CREB in rat brain regions, and the effects of rolipram, a cAMP-specific phosphodiesterase (PDE) inhibitor on the CRE-binding activity by electrophoretic mobility-shift assay (EMSA). A marked age-dependent decrease in the CRE-binding activity was shown in all brain regions examined, especially in the basal forebrain, the striatum and the hippocampus. Furthermore, CRE-binding activities in the basal forebrain of both young-adult and aged rats significantly increased 2 h after rolipram administration (1 mg/kg, i.p.), and the rolipram treatment recovered the decreased CRE-binding activity in the aged rats. The saturation experiment in EMSA also revealed that rolipram reversed the decrease in the maximum CRE-bindings in the basal forebrain with aging. Since the 5' upstream region of the rat choline acetyltransferase (ChAT) gene contains CRE, and ChAT-positive neurons in the basal forebrain project to the frontal cortex and the hippocampus, rolipram may exert its previously reported ameliorating effect on the age-related reductions of ChAT activities in the frontal cortex and the hippocampus by phosphorylating CREB in the basal forebrain with activation of cAMP-dependent protein kinase via inhibition of PDE.
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PMID:Alterations of cAMP response element-binding activity in the aged rat brain in response to administration of rolipram, a cAMP-specific phosphodiesterase inhibitor. 888 54

Chronic morphine administration increases levels of adenylyl cyclase and cAMP-dependent protein kinase (PKA) activity in the locus coeruleus (LC), which contributes to the severalfold activation of LC neurons that occurs during opiate withdrawal. A role for the transcription factor cAMP response element-binding protein (CREB) in mediating the opiate-induced upregulation of the cAMP pathway has been suggested, but direct evidence is lacking. In the present study, we first demonstrated that the morphine-induced increases in adenylyl cyclase and PKA activity in the LC are associated with selective increases in levels of immunoreactivity of types I and VIII adenylyl cyclase and of the catalytic and type II regulatory subunits of PKA. We next used antisense oligonucleotides directed against CREB to study the role of this transcription factor in mediating these effects. Infusion (5 d) of CREB antisense oligonucleotide directly into the LC significantly reduced levels of CREB immunoreactivity. This effect was sequence-specific and not associated with detectable toxicity. CREB antisense oligonucleotide infusions completely blocked the morphine-induced upregulation of type VIII adenylyl cyclase but not of PKA. The infusions also blocked the morphine-induced upregulation of tyrosine hydroxylase but not of Gialpha, two other proteins induced in the LC by chronic morphine treatment. Electrophysiological studies revealed that intra-LC antisense oligonucleotide infusions completely prevented the morphine-induced increase in spontaneous firing rates of LC neurons in brain slices. This blockade was completely reversed by addition of 8-bromo-cAMP (which activates PKA) but not by addition of forskolin (which activates adenylyl cyclase). Intra-LC infusions of CREB antisense oligonucleotide also reduced the development of physical dependence to opiates, based on attenuation of opiate withdrawal. Together, these findings provide the first direct evidence that CREB mediates the morphine-induced upregulation of specific components of the cAMP pathway in the LC that contribute to physical opiate dependence.
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PMID:CREB (cAMP response element-binding protein) in the locus coeruleus: biochemical, physiological, and behavioral evidence for a role in opiate dependence. 931 9

cAMP-dependent protein kinase (PKA) in the nucleus accumbens (NAc) has been implicated in cocaine addiction because (1) cocaine reinforcement is mediated by dopamine receptors that modulate cAMP formation, and (2) repeated exposure to cocaine upregulates the cAMP system in NAc neurons. This study tested PKA involvement in cocaine self-administration and relapse of cocaine-seeking behavior by infusing cAMP analogs that activate or inhibit PKA into the NAc of rats. Bilateral intra-NAc infusions of the PKA inhibitor Rp-cAMPS reduced baseline cocaine self-administration, shifted the dose-response curve for cocaine self-administration to the left, and induced relapse of cocaine-seeking behavior after extinction from cocaine self-administration, consistent with an enhancement of cocaine effects in each paradigm. In contrast, pretreatment with intra-NAc infusions of a PKA activator, Sp-cAMPS or dibutyryl cAMP, increased baseline cocaine self-administration during the second hour of testing and shifted the dose-response curve to the right, consistent with an antagonist-like action. After extinction from cocaine self-administration, similar infusions of Sp-cAMPS induced generalized responding at both drug-paired and inactive levers. As an index of PKA activity in vivo, NAc infusions of Rp-cAMPS reduced basal levels of dopamine-regulated phosphoprotein-32 phosphorylation and blocked amphetamine-induced increases in cAMP response element-binding protein (CREB) phosphorylation. Conversely, NAc infusions of Sp-cAMPS increased phosphorylation of CREB. Together, these results suggest that sustained upregulation of the cAMP system in the NAc after repeated cocaine exposure could underlie tolerance to cocaine reinforcement, whereas acute inhibition of this system may contribute to drug craving and relapse in addicted subjects.
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PMID:Involvement of cAMP-dependent protein kinase in the nucleus accumbens in cocaine self-administration and relapse of cocaine-seeking behavior. 946 9

Here, we report the effects of short exposure to melatonin on the human mt1 (h mt1) melatonin receptor-mediated signaling in Chinese hamster ovary (CHO) cells, and the consequences of an exposure that resembles the physiological pattern of melatonin release on cAMP-mediated signal transduction. Short exposure (10 min) of h mt1 melatonin receptors to melatonin (400 pM) inhibited forskolin-stimulated cAMP formation, cAMP-dependent protein kinase activity, and phosphorylation of the cAMP response element-binding protein. However, treatment of mt1-CHO cells with melatonin in a manner that closely mimics the in vivo activation of melatonin receptors (i.e. 400 pM melatonin for 8 h to mimic darkness) resulted in a supersensitization of the cAMP-dependent signal transduction cascade during the period of withdrawal (i.e. 16 h without melatonin to mimic the light cycle of a diurnal photoperiod). During the period of withdrawal, forskolin induced a time-dependent (1-16 h) increase in cAMP formation (approximately 200% of control cells). This effect of melatonin was dependent on the presence of the h mt1 melatonin receptor, as no potentiation of forskolin-induced cAMP formation was observed in CHO cells transfected only with the neomycin resistance plasmid. The time-dependent increase in forskolin-stimulated cAMP levels resulted in a potentiation of cAMP-dependent protein kinase activity 1 h after withdrawal (approximately 130% of control cells; P < 0.05) and in the number of cells containing the phosphorylated form of cAMP response element-binding protein (approximately 75% of cells at 1 and 16 h compared with 30% in control cells; P < 0.05). An increase in the undissociated state (G alphabetagamma) of Gi proteins may underlie this phenomenon as demonstrated by the increase in pertussis toxin-catalyzed ADP-ribosylation of G proteins (217 +/- 48% of control; P < 0.05) after melatonin withdrawal. This increase in the ribosylation was not due to an up-regulation of Galpha(i) protein, as no significant change in Galpha(i) protein levels occurred at this time. We demonstrated that activation of the h mt1 melatonin receptor in a manner that resembles the physiological pattern of melatonin exposure alters signaling, as potentiation of cAMP-mediated signal transduction events is observed after hormone withdrawal. The CHO cells expressing the human melatonin receptor may provide an in vitro cellular model in which to investigate the putative signaling mechanisms leading to gene regulation by melatonin.
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PMID:Physiological exposure to melatonin supersensitizes the cyclic adenosine 3',5'-monophosphate-dependent signal transduction cascade in Chinese hamster ovary cells expressing the human mt1 melatonin receptor. 964 77

We have analyzed the properties of cAMP response element-binding protein (CREB) in solution with emphasis on dimerization and effects of phosphorylation. Using a purified CREB fusion protein, a novel dye-label technique, and sedimentation equilibrium analysis, we directly and conclusively demonstrate that, unlike Jun and Fos, CREB dimerization is DNA-dependent. CREB exists primarily as a monomer in solution and cooperatively assembles on DNA to form dimers. Sedimentation equilibrium analysis also indicates that dimerization is unaffected by cAMP-dependent protein kinase-phosphorylation or by the symmetry of the cAMP-responsive element binding site. Filter binding assays reveal that CREB binding is unaffected by phosphorylation regardless of the symmetry of the cAMP-responsive element binding site. Our results suggest that structurally similar members of the same bZIP superfamily may differ significantly in their regulation at the level of dimerization.
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PMID:cAMP response element-binding protein monomers cooperatively assemble to form dimers on DNA. 969 27

Gene expression is necessary for the formation and consolidation of long term memory in both invertebrates and vertebrates. Here, we describe the expression and characterization of candidate plasticity gene 16 (cpg16), a protein serine/threonine kinase that was previously isolated from rat hippocampus as a plasticity-related gene. CPG16, when expressed in and purified from bacteria and COS7 cells, was only capable of autophosphorylation and phosphorylation of myelin basic protein but failed to phosphorylate many other peptides and proteins in in vitro phosphorylation assays. Recombinant CPG16, when overexpressed and purified from COS7 cells, had a relatively low level of autophosphorylation activity. This activity was significantly stimulated when cAMP-elevating agents (forskolin, 8-bromo-cAMP) were added to the cells but not by any other extracellular stimuli tested, e.g. serum, phorbol esters, and a calcium ionophore. Although the stimulation of CPG16 activity was inhibited by the cAMP-dependent protein kinase inhibitor H-89, it did not serve as a direct substrate for this kinase. This suggests that CPG16 may be activated by a cAMP-stimulated protein kinase cascade. Immunolocalization studies in COS7 and NIH-3T3 cells showed mostly cytoplasmic localization of CPG16 that turned partially nuclear upon stimulation with 8-bromo-cAMP. Moreover, overexpression of CPG16 seems to partially inhibit cAMP-stimulated activity of the transcription factor CREB (cAMP response element-binding protein), suggesting its involvement in the down-regulation of cAMP-induced transcription. Thus, CPG16 is a protein serine/threonine kinase that may be involved in a novel signaling pathway downstream of cAMP-dependent protein kinase.
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PMID:CPG16, a novel protein serine/threonine kinase downstream of cAMP-dependent protein kinase. 991 91


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